Green Fluorescent Protein (GFP) is a light emitting protein purified from the jellyfish Aequorea victoria. GFP can emit green light by accepting energy transfer from sources that include exogenous blue light and Renilla luciferase catalyzed reactions. The gene for GFP was cloned and its cDNA is a powerful reporter gene in a variety of living systems, including bacteria, fungi, and mammalian tissues. The UV light stimulated GFP fluorescence does not require cofactors and the gene product alone can be sufficient to allow detection of living cells under the light microscope.
By modifying the wild type GFP protein, red-shifted GFP variants with bright emission have also been produced. These variants include EGFP, GFPS65T and RSGF. Recently, GFP was expressed in a human cell line and in vivo. C. Kaether, H. H. Gerdes. Visualization of protein transport along the secretory pathway using green fluorescent protein. FEBS-Lett. 1995; 369:267-71. "Humanized" GFP was synthesized with nucleotide changes that did not change the amino acid sequences with one exception.
Renilla luciferase is an enzyme purified from Renilla reniformis. The enzyme catalyzes the oxidative decarboxylation of coelenterazine in the presence of oxygen to produce blue light with an emission wavelength maximum of 478 nm. In Renilla reniformis cells, however, this reaction is shifted toward the green with a wavelength maximum of 510 nm due to an energy transfer to a Green Fluorescent Protein.
The gene for Renilla luciferase (ruc) was cloned and its cDNA was shown to be useful as a reporter gene in various living systems. D. C. Prasher, V. K. Eckenrode, W. W. Ward, F. G. Prendergast, M. J. Cormier. Primary structure of the Aequorea victoria green-fluorescent protein. Gene 1992; 111:229-33. By providing appropriate promoters to the cDNA as gene cassettes, the gene was expressed in bacteria, transformed plant cells, and mammalian cells. The high efficiency of Renilla luciferase is a useful trait as a marker enzyme for gene expression studies.
Given the properties of GFP and Renilla luciferase, it would be useful to have a single protein combining the functions of both Renilla luciferase enzymes and GFP to monitor gene expression quantitatively by UV light excitation or qualitatively by enzyme activity measurements.